A Newly Identified Peripheral Duplex Anchors and Stabilizes the MALAT1 Triplex

The accumulation of the 8-kb oncogenic long noncoding MALAT1 RNA in cells is dependent on the presence of a protective triple helix structure at the 3' terminus. While recent studies have examined the functional importance of numerous base triples within the triplex and its immediately adjacent base pairs, the functional importance of peripheral duplex elements has not been thoroughly investigated. To investigate the functional importance of a peripheral linker region that was previously described as unstructured, we employed a variety of assays including thermal melting, protection from exonucleolytic degradation by RNase R, small-angle X-ray scattering, biochemical ligation and binding assays, and computational modeling. Our results demonstrate the presence of a duplex within this linker that enhances the functional stability of the triplex in vitro, despite its location more than 40 Å from the 3' terminus. We present a full-length model of the MALAT1 triple helix-containing RNA having an extended rod-like structure and comprising 33 layers of coaxial stacking interactions. Taken together with recent research on a homologous triplex, our results demonstrate that peripheral elements anchor and stabilize triplexes in vitro. Such peripheral elements may also contribute to the formation and stability of some triple helices in vivo.